CONTROLLED

DELIVERY OF
VACCINE
CONTROLLED DELIVERY
OF VACCINE USING
BIODEGRABLE
SUBSTANCE
 ORIGIN OF VACCINE
 Smallpox was the first disease which are
inoculated with other type of type
infections which was believed to be started
in India and China before 200BC
 Scientists who are involved are

1) Lady Mary Worltey Montague

2) Edward Jenner

3) Sarah Nelmes

4) James Phipps
 DEVELOPMENT OF NEW
VACCINE
1998 first vaccine for
rotavirus was developed;
21st century in 2006
human papillomavirus was
developed for cancer.
DEFINATION OF VACCINE
 Vaccine which are
suspensions of killed, lived,
or attenuated (having
weakened virulence)
cultures of micro-organism
are used as antigen to
produce immunity against
infection due to the
particular microorganism
 Example→ typhoid fever
vaccine consists of killed
cell salmonella typhii
CHOLREA
 TYPE OF VACCINE
 INACTIVATED VACCINE- Microorganism that has
been killed chemicals or heat. This type of
vaccine has less immune response due to which
booster doses require.
 Example – vaccine against flu, cholera

2) Live attenuated – These are live microorganism

that has been cultivated under condition which
disables their virulent properties. They have
durable immunological reponse
Example →yellow fever, measles, rubella and
mumps
CLASSIFICATION OF COMMON
USED VACCINE
DISEASE TYPE OF
VACCINE
Tuberculosis Attenuated
Cholera Inactivated
Polio Attenuated
Influenza Inactivated
Diphtheria Inactivated
exotoxin
H influenza type Polysaccharide+
IMMUNIZATION SCHEDULE FOR
CHILDREN IN INDIA
AGE OF CHILD VACCINE

1& ½ BCG, DPT*1 AND

PILIO*1
2&½ DPT*2 & PILIO*2

3&½ DPT*3 & PILIO*3

9 months Measles

Below 16 and 24 DPT*4 & PILIO*4

months
DEVELOPMENT AND DESIGNING
OF NEW GENERATION VACCINE
1) Multivalent subunit 5) Antigen vaccine
vaccine (a) Recombinant
(a)Small matrix antibody vaccines
antigen complexes 6) Vector vaccines
(b) Liposomes (d) Recombinant vector
2)Purified vaccine
macromolecules 7) Anti idiotype vaccines
3)Synthetic peptides as 8) Targeted immune
vaccine stimulants
4) Immunoadhesions
 OBJECTIVE OF THE VACCINE
DELIVERY
 To elicit a protective immune response of
sufficiently long duration, from a single-
contact immunization
 Potentiate the immune response to
vaccine without manifesting any adverse
effects on the body
 Incorporate many in a single formulation
deliver even those vaccines through the
oral route that currently need to be given
parenterally.
 INDUCTION OF IMMUNE
RESPONSE
 Vaccine is carried out to protect the
individual from by priming the immune
system to resist the infecting agent.
 This resistance can be offered by effectors
molecules antibodies, cytokines,
complement etc or effectors cell (cell
mediated immunity).
 Immunization also results in the system
developing a “memory” of exposure to
antigen present on the pathogen
 HOW IMMUNE RESPONSE IS
GENERATED ?
 Immune response
1) Cell type immune response
a) Non specific cell type
i) Phagocytes
ii) Auxiliary cell
b) Targeted specific cell type
i) B cell
ii) T cell
2) Effector response
 MECHANISM ACTION OF THE
CELL TYPE IMMUNE RESPONSE
 Phagocytes and auxiliary are
first line defence against
infection
 Inflammatory substance
released from auxiliary cell on
encounter with the foreigen
material, stimulate
phagocytes and attract from
the site of infection.
 These cell get engulf the
material upon the phagocytes
 Fragments of proteins
produce are essential for
generation of a T cell
 While performing the function
macrophage act as antigen
presenting cell (APC)
 The carry metabolic
organelles to kill micro-
organism by generating the
free radical
 MECHANISM OF
IMMUNOGENCITY BY CARRIER
MOLECULES
 The coupling of small peptides to carrier proteins
increase the molecular mass of peptide and there
by improves the uptake by APC

 Biological half of small peptides may be

increased via the linkage to carrier protein

 If the synthetic peptides itself is immunogenic

(as in the case of peptides having B cell and T
cell epitopes ), the carriers proteins merely
functions as a polymeric delivery system.
 MECHANISM OF
IMMUNOGENICITY VIE CARRIER
MOLECULES
 Commonly used
carrier are
2) Keyholes limpet
hemocyin (KLH) and
sperm whale
myoglobin (SWM)
3) Albumins
(ovalbumins)
4) Fig will show the
attachment of hapten
to carrier molecules
 EFFECTOR RESPONSE
 Effectors response of the T cell consist of secreting
specialized substance called cytokines.
 Cytokines are divided into two broad class
3) Helper T cell help B & T cell in variety of ways.
4) Cytokines or “killer” T cell make the cytokines that killed
the infected cells
Antibodies produce during this process are help in the in
binding with invading microorganism and facilitating the
pathogens bearing the Ag phagocytes.
T cell are divided into TH1 which are primary involved in
cell mediated immunity and TH 2 with the production of
antibodies
►T cell are divided
into TH1 which are
primary in cell
mediated and TH 2
with the production
►TH 1 secrete
gamma interferon
TNF
►TH2 secrete IL-2,
IL-5,IL-6, IL-10
 CELL MEDIATED IMMUNITY
 On the exposure to the appropriate antigen T
lymphocytes of lymphoid tissue proliferated to
form activated T cell which are of three type
1) Helper T cell – release lymphokines which
stimulate growth and differentiate of the
activated B cell to from plasma cell
2)Cytotoxic T cell which bind tightly to specific
binding antigen which then form secrete pore and
form round role in the membrane
3) Suppresser T cell tthey are believed to inhibit the
conversion of B cell into plasma cell
 DESIGING RATIONALE AND
REQUIRMENT OF AN EFFICIENT
VACCINE DELIVERY
 Particulate
Antigen
 Booster Doses

 Adjuvant – commonly used is

Calcium phosphate
● Antigen Stability
 PARTICULATE ANTIGEN
 Uptake is generally take by phagocytes which are
important for eliciting an immune response: the
vaccine formulation should be amenable to
phagocytosis.
 Soluble material which can be moved about
anywhere is taken up by the pinocytosis. which is
10 times less efficiency than phagocytosis
 The efficiency of uptake can make the
formulation in a particulate form.
 Finally particulate matter is important signal for
phagocytic cells to home towards the site
 BOOSTER DOSES
 Vaccine that are required to be given in
the divided doses to produce the more
efficiency
 High doses tolerance is another curious
phenomenon which occur if size of the
doses is too high which rendered the
antigen and no response is produced
 Efficiency of the divide doses can be
sought with the reference to cellular
events that takes place in the
immunization
 ADJUVANTS
 Vaccine formulation is usually administered with
the special additive called “adjuvant “ special
when the vaccine is in the form of killed
pathogens or isolated protein instead of live
attenuated organism”
 “ A VACCINE DELIVERY SYSTEM IS ALSO
EXPECTED TO HAVE SUFFICIENT
ADJUVANTTICITY TO POTENTIATE THE
REPONSE TO DELIVERY ANTIGEN.”
 Commonly used adjuvant are calcium phosphate
which act as adjuvant and improved safety profile
and enhance immune system stimulation
 PRODUCTION OF CAP
♦The formulation of CAP nanoparticles is easily tailored for
each antigen.
 The manufacturing process is quick and requires only
simple equipment, water for injection, and inorganic
salts.
 The CAP nanoparticle have exceptional protein loading
capacity: about 20%(w/w) if antigen is coated on the
surfaces only and about 50% and greater for internal
“core loading”.
 Batch-to batch consistency is excellent.
 Long-term storage (out to one year) resulted in no
changes in particles size, ph and surface morphology
 ANTIGEN STABILITY
 Antigen stability – antigen used for
immunization is subjected to
environmental stress such as temperature
ph or non-aqueous me3dium which may
cause the change confirmation.
 This may result the production of epitopes
which are meant for generate the B cell
response
 Requirement for maintaining the stability
of the antigen are follows – low
temperatures but not freezing, physilogical
ph and aqueous environment
 PREPARATION OF
MICROSPHERE IN CONTROLLED
DELIVERY
 The properties for preparation of
microsphere are
1) optimal antigen loading
2) size of microsphere
3) minimum wastage of material
4) within batch uniformity and inter-batch
reproducibility
5) minimum exposures of Ag to denaturing
condition
6) For parentrally products microsphere
need to be sterile
 POLY ( LACTIDE-CO-GLYCIDE)
(PLGA)
 This type of the microsphere which is used for the
controlled delivery system of peptides, native and
synthetic proteins and lately.
 PLGA microsphere are composed of a spherical
shaped polymeric matrix ranging in diameter
from 1 to 250 micrometere.
 Many factors are important to formulate
 i) Ability to release the entrapped substance

ii) Particle size

iii) Stability and safety which is related to in vivo
polymer degration
 Antigen are physically entrapped into
microsphere inject able solid polymeric matrix
 The combination of diffusion through pores and of
polymer matrix biodegradable allows the control
of antigen release rates
 The biodegrading rates of polymer depends on its
molecular weight
 After hydrolysis degraded products form
monomer of lactide and glycolide which are
eliminated by Krebs cycle as carbon dioxide &
urine.
 During the process, the encapsulated antigen are
release which can varies from hours to months
depending upon the polymer combination
 Microsphere contd
 Microsphere diameter plays an important role in
interaction with phagocytic cells
 Particle smaller than the 10 micrometer
phagocytosed faster by the macrophage which
cause recruitment to the site of administration
after subcutaneous injection.
 Particle more than 10 micrometer act as depot
releasing the antigen.
 Polymer selection also plays an important role in
the manufactures because it critically influence
their rate of biodegrading
 By changing the Contd
homopolymer ratio
different physicochemical
 compositions can
influence its degradability
and permeability
 These parameter can be
easily monitored using
different polymer and
copolymer
 Amorphous polymer are
more permeable than
PLGA
 The combination of
particles with different
compositions in the same
formulation has permitted
 MICROSPHERE PERPARATION
 This can be done by two methods
1) Simple emulsification (O\W)
2) Multiple water–in-oil-water (W\O\W)
♦ the choice of method depends on the physical and chemical
characteristics of antigen, permitting the matrix entrapment of both
lipophilic and hydrophilic
♦ in the first case an emulsion is formed by dissolving the antigen in an
organic solvent immiscible in water (methlyene chlorides )
containing the polymer , under strong mechanical agitation .
this emulsion can be stabilized by added surfactants to the aqueous
phase.
The solvent is eliminated by the evaporation at room temperatures,
followed by the washed and freezing drying
 CONTD
 The multiple emulsion method involves water-in-oil-in-
water emulsification
 The inner aqueous phase containing the hydrophilic
substance is obtained after emulsification with the
immiscible organic solvent containing polymer under strong
mechanical agitation.
 This emulsion is stabilized by the addition of aqueous
solution containing a surfactants (e.g. poly vinyl alcohol )
and is further homogenized to produce a W\O\W double
emulsion.
 This double emulsion is gently stirred with homogenizer
room temperatures for solvent evaporation
 The microsphere are collected by the centrifugation,
washed with water and freeze dried.
 the mixture of hydrphilic and lipophilic molecules can be
successfully entrapped by this method
 PGLA MICROSPHERE
BIOCOMPATIBLITY
 Polymer microspheres have attracted much attention because of their
biocompatible characteristics the phagocytosis of biodegradable and
no biodegradable particles has reported to depend on their size, surface
charge and hydrophobicity.
♦ After subcutaneous administration, PLGA microsphere ranging in the
diameter from 1 to 10 micrometer is readily phagocytosed by
macrophages recruited to the site of injection, thereby providing an
intracellular delivery of the antigen.
♦This mechanism may enhance antibody response and consequently
decreases the required antigen dose. On the other hand, particles larger
than 10 micrometer in diameter would remain as a depot at the site of
injection providing a sustained release of antigen.
 Improving their hydrophobicity can increase phagocytosis of the particles,
whereas microsphere preparation with different compositions of PLA and
 PLGA do alter the extent of phagocytosis.
 ROUTE OF IMMUNIZATION
 The oral route of delivery is the most acceptable in terms of patient
compliance.
 the antigen entering the body through the oral route include items of
food, and are “tolergenic”.
 Lymphoid tissue associated with the digestive tract is located
primarily in Peyer’s patches in the small intestine and the material
that targets to Peyer’s patches often evokes an immune response.
 This response is often in a such type as protect against pathogens

entering the body by invading mucosal tissue if the response against

microbes invading by non-mucosal routes is desired after oral
immunization, the delivery system must be capable of transporting
the Ag to deep sealed lymphoid tissue instead of releasing it to the
mucosa-associated lymphoid tissue.
 Systemic routes of immunization should seek a route that ensures

maximal exposures of the Ag to immunocytes.

 The usual choices are intramuscular (i.m.) subcutaneous or

interdermal immunization.
As th e site deliv ery mo ves fr om
deep tis sues towa rd s su perfi cial,
the d eli very syste m is e xpose d to
a la rg er n umb er o f fi rst l in es
defe nce cell (i. m, i.d ) at th e same
time , the perio d o f re sid ence of
the d eli very syste m in the b ody
decrease a s the sit e o f deliv ery
moves t owa rds su perfic ia l tiss ue .
Route of immunization also takes place
by two way 1) Systemic immunization
2) Mucosal immunization
 CONCLUSION
 PLGA microsphere with an incorporated
antigen represents a good antigen delivery
system for both cellular and humoral
response.
 The easy manufactures of microsphere
and the possibility of administration by
different routes offer the additional
advantage of their use as a
pharmaceutically acceptable adjuvant for
vaccines.
 INSTRUMENTAL METHOD OF
PERPARATION
 This method include
2) Spray drying
3) Air suspension coating
4) Press grinding
5) Coaceration-Phase separation
i) Emulsification methods
a) Oil-in-oil emulsion
b) Oil-in-water emulsion
c) Multiple emulsion
5) Solvent extraction
6) Rotary evaporations
 SPRAYING DRYING
 In this method spray dried protein
is suspended in a solution of the
polymer in an organic solvent
(methylene chloride
ortetrahydronfuran).
1)This is then pumped into spray drier
by a peristaltic pump.
2) Dry air at high pressure and inlet
air temperature of 37ºC atomizes
the air suspension, and the
polymer forms a matrix entrapping
the protein as the solvent
evaporates from the droplets.
 Spherical particles of the desired
size range and loading can thus
prepared.

DISADVANTAGE OF THIS
METHOD
 This method is relatively
large batch size required for
processing in view of the
limitation of the
availableequipment.
♦ Even small volumes spray
driers are not designed to
handle volumes of the order of
a few ml, which would be
sufficient to produce
thousands of doses.
 This size dispersion of
microsphere produced by this
method is also exposed to be
large.
 EVALUATION OF VACCINE
LAODED MICROSPHERE
 The evalulation factors involve
1)Praticle size
2) Antigen loading
3) Structural integrity of
encapsulated antigen
4)Studies on the antigen release in
vitro
5) Antigencity of the preparation
Praticle size
 Particles size can be
easily determine using
light microscopy or in the
greater detail by scanning
the electron microscopy
(SEM) which shows the
scanning electron
microphotography of
abatch of the
microsphere.
 SEM also helps in
accessing surface
morphology of the
microsphere.
 Standards equipment for
micromerities such as
Coulter Counter or laser
 Studies on the antigen release in
vitro
 In vitro Ag release studies
have served as quality
control parameters as
well as index of the
duration of immune
response.
 These help to understand
the nature of release and
provide significant insight
into the formulation
variables
 By dispersing the
microsphere in a suitable
buffer in a vial and
subjecting it to stirring at
37ºc, over the period of
time.
 NOVEL DRUG SYSTEM
Novel drug system include following point
1)Particulate delivery system
i) Liposome
ii) Emulsion
iii) Micro sphere
2)Cochleates
3)Mucoadhesive polymer
4)DNA vaccination
5)Transgenic plants an edible immunogen concept